The effects of hormone-mediated PI3K/AKT signaling on spermatogenesis in Sertoli cells

The phosphoinositide-3-kinase/Akt(PI3K/AKT)signaling pathway is crucial for Sertoli cell development and completing spermatogenesis.Its main role is to promote proliferation and inhibit apoptosis.Many factors activate the PI3K/AKT pathway,like hormones,such as follicle stimulating hormone(FSH),androgen,estrogen,insulin to name a few.Many of these factors have receptors inside or on the surface of Sertoli cells(SCs).This review summarizes how these hormones directly regulate the PI3K/AKT signaling pathway in SCs,which in turn affects SC proliferation and differentiation.Further,hormone-mediated PI3K/AKT signaling also stimulates SC secretion,which is essential for germ cell development,suggesting an indirect role of PI3K/AKT signaling during spermatogenesis.These functions include promoting spermatogonia proliferation and differentiation,meiosis of spermatocytes,sperm maturation,and their release.This review also provides potential hints for clinically treating male infertility issues like cryptorchidism and Sertoli cell-only syndrome.

Rikkunshito increases peripheral incretin-hormone levels in humans and rats

BACKGROUND It was reported that rikkunshito(TJ-43)improved the cisplatin-induced decreases in the active form of ghrelin in plasma;however,other effects on gastrointestinal hormones have not been investigated.AIM To investigate the effects of TJ-43 on peripheral levels of incretin hormones,including gastric inhibitory polypeptide(GIP)and glucagon-like polypeptide-1(GLP-1),in humans and rats.METHODS Patients were divided into two groups,namely patients who received TJ-43 immediately following surgery[TJ-43(+)group]and those who received TJ-43 on postoperative day 21[TJ-43(-)group],and the plasma levels of active GIP and active GLP-1 were assessed.In animal experiments,rats were treated with TJ-43[rat(r)TJ-43(+)group]or without[rTJ-43(−)group]by gavage for 4 wk,and the plasma active GIP and active GLP-1 levels were measured.The expression of incretin hormones in the gastrointestinal tract and insulin in the pancreas were investigated by immunohistochemistry.Furthermore,the cyclic adenosine monophosphate activities were assessed in pancreatic tissues from rats treated with or without TJ-43 in vivo,and the blood glucose levels and plasma insulin levels were measured in rats treated with or without TJ-43 in oral glucose tolerance tests.RESULTS In humans,the active incretin hormone levels increased,and values were significantly greater in the TJ-43(+)group compared those in the TJ-43(-)group.In rats,the plasma active incretin levels significantly increased in the rTJ-43(+)group compared with those in the rTJ-43(-)group.GIP and GLP-1 expressions were enhanced by TJ-43 treatment.Moreover,plasma insulin levels increased and blood glucose levels were blunted in the rTJ-43(+)group.CONCLUSION The results show that TJ-43 may be beneficial for patients who undergo pancreatic surgery.

Gastrointestinal hormone abnormalities and G and D cells in functional dyspepsia patients with gastric dysmotility

AIM: To investigate the relationship between gastric dysmotility,gastrointestinal hormone abnormalities, and neuroendocrine cells in gastrointestinal mucosa in patients with functional dyspepsia (FD).METHODS: Gastric emptying was assessed with solid radiopaque markers in 54 FD patients, and the patients were divided into two groups according to the results, one with delayed gastric emptying and the other with normal gastric emptying. Seventeen healthy volunteers acted as normal controls. Fasting and postprandial plasma levels and gastroduodenal mucosal levels of gastrointestinal hormones gastrin, somatostatin (SS) and neurotensin (NT)were measured by radioimmunoassay in all the subjects.G cells (gastrin-producing cells) and D cells (SS-producing cells) in gastric antral mucosa were immunostained with rabbit anti-gastrin polyclonal antibody and rabbit anti-SS polyclonal antibody, respectively, and analyzed quantitatively by computerized image analysis.RESULTS: The postprandial plasma gastrin levels, the fasting and postprandial plasma levels and the gastric and duodenal mucosal levels of NT were significantly higher in the FD patients with delayed gastric emptying than in those with normal gastric emptying and normal controls. The number and gray value of G and D cells and the G cell/D cell number ratio did not differ significantly between normal controls and the FD patients with or without delayed gastric emptying.CONCLUSION: Our findings suggest that the abnormalities of gastrin and NT may play a role in the pathophysiology of gastric dysmotility in FD patients, and the abnormality of postprandial plasma gastrin levels in FD patients with delayed gastric emptying is not related to the changes both in the number and gray value of G cells and in the G cell/D cell number ratio in gastric antral mucosa.

Gonadotrophin-releasing hormone-Ⅰ and -Ⅱ stimulate steroidogenesis in prepubertal murine Leydig cells in vitro

Aim： To study the effect and mechanism of gonadotrophin-releasing hormone （GnRH） on murine Leydig cell steroidogenesis. Methods： Purified murine Leydig cells were treated with GnRH-Ⅰ and -Ⅱ agonists, and testosterone production and steroidogenic enzyme expressions were determined. Results： GnRH-Ⅰ and -Ⅱ agonists significantly stimulated murine Leydig cell steroidogenesis 60%-80% in a dose- and time-dependent manner （P 〈 0.05）. The mRNA expressions of steroidogenic acute regulatory （STAR） protein, P450scc, 3β-hydroxysteroid dehydrogenase （HSD）, but not 17β-hydroxylase or 17β-HSD, were significantly stimulated by both GnRH agonists with a 1.5- to 3-fold increase （P 〈 0.05）. However, only 3β-HSD protein expression was induced by both GnRH agonists, with a 1.6- to 2-fold increase （P 〈 0.05）. Conclusion： GnRH directly stimulated murine Leydig cell steroidogenesis by activating 3β-HSD enzyme expression.

Effects of luteinizing hormone and androgen on the development of rat progenitor Leydig cells in vitro and in vivo

Progenitor Leydig cells are derived from stem cells. The proliferation and differentiation of progenitor Leydig cells significantly contributes to Leydig cell number during puberty. However, the regulation of these processes remains unclear. The objective of the present study was to determine whether luteinizing hormone （LH） or androgen contributes to the proliferation and differentiation of progenitor Leydig cells. Fourteen-day-old male Sprague-Dawley rats were treated for 7 days with NalGlu, which is a gonadotropin- releasing hormone antagonist, to reduce the secretion of LH in the pituitary and thus, androgen in the testis. Rats were co-administered with LH or 7a-methyl-nortestosterone （MENT）, which is an androgen resistant to metabolism by 5a-reductase 1 in progenitor Leydig cells, and the subsequent effects of LH or androgen were measured. 3H-Thymidine was also intravenously injected into rats to study thymidine incorporation in progenitor Leydig cells. Progenitor Leydig cells were examined. NalGlu administration reduced progenitor Leydig cell proliferation by 83%. In addition, LH or MENT treatment restored Leydig cell proliferative capacity to 73% or 50% of control, respectively. The messenger RNA levels of proliferation-related genes were measured using real-time PCR. The expression levels of Igfl, Lifr, Pdgfra, Bcl2, Ccnd3and Pcnawere upregulated by MENT, and those of Pdgfra, Ccnd3and Pcnawere upregulated by LH. Both LH and MENT stimulated the differentiation of progenitor Leydig cells in vitro. We concluded that both LH and MENT were involved in regulating the development of progenitor Leydig cells.

Maturation, proliferation and apoptosis of seminal tubule cells at puberty after administration of estradiol, follicle stimulating hormone or both

Aim： To assess proliferative and apoptotic potential of the seminiferous epithelium cells in relation to Sertoli cell maturation in newborn rats under the influence of estradiol, follicle stimulating hormone （FSH） or both agents given together. Methods： From postnatal day （PND） 5 to 15 male rats were daily injected with 12.5 μg of 1713-estradiol benzoate （EB） or 7.5 IU of human purified FSH （hFSH） or EB ＋ hFSH or solvents （control）. On postnatal day 16, autopsy was performed. Sertoli cell maturation/function was assessed by morphometry. Proliferation of the seminiferous epithelium cells was quantitatively evaluated using immunohistochemical labeling against proliferating cell nuclear antigen and apoptosis using the TUN-EL method. Results： Although EB inhibited Sertoli cell maturation and hFSH was not effective, a pronounced acceleration of Sertoli cell maturation occurred after EB ＋ hFSH. Whereas hFSH stimulated Sertoli cell proliferation, EB or EB ＋ hFSH inhibited Sertoli cell proliferation. All treatments significantly stimulated germ cell proliferation. Apoptosis of Sertoli cells increased 9-fold and germ cells 2-fold after EB, and was not affected by hFSH but was inhibited after EB ＋ hFSH. Conclusion： At puberty, estradiol inhibits Sertoli cell maturation, increases Sertoli and germ cell apoptosis but stimulates germ cell proliferation. Estradiol in synergism with FSH, but neither of the hormones alone, accelerates Sertoli cell maturation associated with an increase in germ cell survival. Estradiol and FSH cooperate to induce seminal tubule maturation and trigger first spermatogenesis.

Impact of parathyroid hormone on bone marrow-derived stem cell mobilization and migration

Parathyroid hormone(PTH) is well-known as the principal regulator of calcium homeostasis in the human body and controls bone metabolism via actions on the survival and activation of osteoblasts. The intermittent administration of PTH has been shown to stimulate bone production in mice and men and therefore PTH administration has been recently approved for the treatment of osteoporosis. Besides to its physiological role in bone remodelling PTH has been demonstrated to influence and expand the bone marrow stem cell niche where hematopoietic stem cells, capable of both self-renewal and differentiation, reside. Moreover, intermittent PTH treatment is capable to induce mobilization of progenitor cells from the bone marrow into the bloodstream. This novel function of PTH on modulating the activity of the stem cell niche in the bone marrow as well as on mobilization and regeneration of bone marrow-derived stem cells offers new therapeutic options in bone marrow and stem cell transplantation as well as in the field of ischemic disorders.

Effects of Xiangsha Liujunzi decoction drug serum on gastric antrum smooth muscle cells from rats with functional dyspepsia by regulating gastrointestinal hormones

Objective:To observe the effect and mechanism of Xiangsha Liujunzi decoction(XSLJZD)drug serum on gastric antrum smooth muscle cells(SMCs)in rats with functional dyspepsia(FD).Methods:Gastric antrum SMCs from rats with FD were isolated,cultured,and then divided into six groups as follows:control,model,domperidone,low-dose XSLJZD(LXSLJZD),medium-dose XSLJZD(MXSLJZD),and high-dose XSLJZD(HXSLJZD).Each group was administered the corresponding drug serum for intervention.Drug serum intervention conditions and proliferative activity of SMCs were tested by cholecystokinin octapeptide.Ghrelin,gastrin,somatostatin,and substance P(SP)levels were measured by ELISA.Somatostatin and SP mRNA expression was measured by real-time PCR.Results:A concentration of 10%drug serum for 24 h was decided to be the best intervention condition for later study.The mean optical density value in the model group was lower than that in the control group(P紏.001).Optical density values in the domperidone and HXSLJZD groups were higher than those in the model group(P?.025,P?.032,respectively).Gastrin,SP,and ghrelin levels in the model group were lower(P?.007,P?.037,P?.005,respectively),but somatostatin levels were higher,compared with those in the control group(P?.031).Gastrin,SP,and ghrelin levels in the domperidone,MXSLJZD,and HXSLJZD groups were higher than those in the model group(all P<.05).Somatostatin levels in the four drug-treated groups were lower than those in the model group(P?.002,P?.007,P?.001,P?.009,respectively).SP mRNA levels in the model group were lower than those in the control,domperidone,MXSLJZD,and HXSLJZD groups(P?.037 P?.016,P?.025,P?.002,respectively).Somatostatin mRNA levels in the model group were higher than those in the control and MXSLJZD groups(P紏.042,P紏.035).Conclusions:XSLJZD and domperidone drug serum effectively promote proliferative activity of gastric antrum SMCs in an FD model.The mechanism of this activity may be regulated by gastrointestinal hormones.

Anti-Müllerian Hormone and Its Utility in Cattle Reproduction

Reproductive biotechnologies offer us greater possibilities to improve animal genetics. However, the success of these depends on different factors such as the proper selection of the donor female. For this reason, endocrine markers have been used to evaluate the ovarian reserve, which allow a successful selection of donors. Recent research has shown, among other things, that concentrations greater than 0.130 ng/mL of anti-Mullerian hormone (also known as Muller-inhibiting substance, which is a member of the transforming growth factor beta superfamily of growth and differentiation factors) are related to donors of more than fifteen transferable embryos. Therefore, this review describes studies showing that the measurement of anti-Müllerian hormone concentrations, before superovulation programs, reduces the costs per embryo produced.

EFFECT OF INTERLEUKIN-1β ON GROWTH HORMONE GENE EXPRESSION AND ITS POSSIBLE MOLECULAR MECHANISM IN RAT MtT/S SOMATOTROPH CELLS

Objective To elucidate the effect of interleukin-1β （IL- 1β） on human growth hormone （hGH） gene expression in a rat somatotropic pituitary cell line MtT/S. Methods Stably transfected MtT/S cells were firstly established by transfecting 484-Lucl plasmid which contained hGH gene promoter --484 to ＋30 bp and luciferase reporter gene. The effect of IL-1β on hGH gene expression was determined by assaying the luciferase activities. RT-PCR method was also used to determine whether IL-1 recepor mRNA was expressed in MtT/S cells. Results The 10^3 U/mL IL-1β stimulated secretion and synthesis of GH, and promoted the 5＇-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.38 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase （MAPKK/MEK） inhibitor PD98059 （40 μmol/L） and p38 mitogen-activated protein kinase （MAPK） inhibitor SB203580 （5 μmol/L） completely blocked the stimulatory effect of IL-1μ, and phosphatidylinositol-3-kinase （PI3-K） inhibitor LY294002 partly abolished the effect of IL-1μ. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit- 1 nor inhibition of Pit- 1 expression affected induction of hGH promoter activity by IL-1μ. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-1β, and results showed that the stimulatory effect of IL-1β was abolished following deletion of the --196 to -- 132 bp fragment. Conclusions IL-1β promotes GH secretion and synthesis in rat MtT/S somatotroph cells. The stimulatory effect of IL-1β on hGH gene promoter appears to require the activation of MEK, p38 MAPK, PI3-K, and a fragment of promoter sequence that spans the -196 to -132 bp of the gene, but it may be unlinked with Pit-1 protein.

Effect of recombinant human growth hormone and interferon gamma on hepatic collagen synthesis and proliferation of hepatic stellate cells in cirrhotic rats

BACKGROUND: Fibrosis plays a key role in the development of liver cirrhosis. In this study, we investigated the effect of growth hormone and interferon gamma on hepatic collagen synthesis and the proliferation of hepatic stellate cells in a cirrhotic rat model. METHODS: Cirrhosis was induced in rats using carbon tetrachloride. Rats were simultaneously treated with daily subcutaneous injections of recombinant human growth hormone or interferon gamma combined with recombinant human growth hormone. The control group was given saline. The relative content of type I and type IV collagen was assessed by indirect immunofluorescence analysis. Activated hepatic stellate cells were prepared from cirrhotic rats. The 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) method was used to assess the effects of recombinant human growth hormone and interferon gamma on these cells in vitro. RESULTS: Both qualitative and quantitative analysis showed that type I and type IV collagen secretion increased with time after recombinant human growth hormone administration and was significantly higher than control and recombinant human growth hormone combined with interferon gamma administration. In vitro, recombinant human growth hormone significantly stimulated hepatic stellate cell proliferation in a concentration-dependent manner (10 -3 -10 -1 mg/100 μL), andinterferon gamma (10 -2 -10 -1 μg/100 μL) significantly inhibited their growth compared to the control group. Interferon gamma combined with recombinant human growth hormone eliminated this growth-promoting effect to a certain degree in a concentration-dependent manner (10 -1 μg/100 μL, P<0.05, 10 -2 -10 -3 μg/100 μL, P>0.05) and a time-dependent manner (P<0.05). CONCLUSIONS: Recombinant human growth hormone increased collagen secretion in cirrhotic rats in vivo and promoted the proliferation of hepatic stellate cells from cirrhotic rats in vitro. It is possible that concurrent interferon gamma therapy can offset these side-effects of recombinant human growth hormone.

MEK AND p38 MAPK-DEPENDANT PATHWAYS ARE INVOLOVED IN THE POSITIVE EFFECT OF INTERLEUKIN-6 ON HUMAN GROWTH HORMONE GENE EXPRESSION IN RAT MtT/S SOMATOTROPH CELLS

Objective To investigate the effect of interleukin-6(IL-6)on the human growth hormone(hGH)gene expression in a rat somatotropic pituitary cell line MtT/S.Methods The plasmids containing various lengths of hGH gene 5'-promoter fragments were constructed.Stably transfected MtT/S cells were created by cotransfecting the above plasmids and pcDNA3.1(+)with DMRIE-C transfection reagent.After the administration of these cells with IL-6 and/or various inhibitors of signaling transduction pathways,the luciferase activities in MtT/S cells lysis were assayed to demonstrate the effects of IL-6 on hGH gene promoter activity and possibly involved mechanism.Results The 103 U/mL IL-6 stimulated GH secretion and synthesis,and promoted the 5'-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.69 times above the control.Among inhibitors of signaling transduction pathways,mitogen-activated protein kinase kinase(MAPKK/MEK)inhibitor PD98059(40 μmol/L)and p38 mitogen-activated protein kinase(MAPK)inhibitor SB203580(5 μmol/L)completely blocked the stimulatory effect of IL-6.Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells.Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected IL-6 induction of hGH promoter activity.A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-6.The results showed that the stimulatory effect of IL-6 was abolished following deletion of the-196 to-132 bp fragment.Conclusions IL-6 promotes GH secretion and synthesis by rat MtT/S somatotroph cells.The stimulatory effect of IL-6 on hGH gene promoter appears to require the activation of MEK and p38 MAPK,and a fragment of promoter sequence that spans the-196 to-132 bp of the gene,but may be unlinked with Pit-1 protein.

Effect of parathyroid hormone on apoptosis of human medullary thyroid carcinoma cells

Objective The aim of the study was to investigate the effect of parathyroid hormone(PTH) on the apoptosis of human medullary thyroid carcinoma(MTC) cells.Methods In vitro cultured medullary thyroid carcinoma cell lines were treated with parathyroid hormone and parathyroid hormone receptor-monoclonal antibody,and the apoptosis of cells was detected by flow cytometry.Results The cell morphology changed significantly after treatment based on the observation using the inverted phase-contrast microscope.Various concentrations of parathyroid hormone and parathyroid hormone receptor-monoclonal antibody effectively induced apoptosis in a time-and concentrationdependent manner.When the concentration of parathyroid hormone was 2.0 μmol/L and that of parathyroid hormone receptor-monoclonal antibody was 1.0 μmol/L,the apoptotic rate was 13.24% and 20.78%,respectively,representing a statistically significant difference from that of the control cells(P < 0.05).Conclusion PTH plays a role in inducing apoptosis of human MTC cells.

Inhibition effects of parathyroid hormone on human medullary thyroid carcinoma cells

Objective： The purpose of the study was to investigate the effects of parathyroid hormone and parathyroid hormone receptor monoclonal antibody on in vitro growth and proliferation of human medullary thyroid carcinoma cell lines. Methods： The medullary thyroid carcinoma cell line was cultured in vitro, with parathyroid hormone and parathyroid hormone receptor monoclonal antibody treatment intervention, the growth of the cells was observed under an inverted contrast micro scope, the MTT assay was used to detect the cell growth inhibition rate. Results： Under the inverted contrast microscope, the cells changed significantly, the parathyroid hormone and parathyroid hormone receptor monoclonal antibodies can effectively inhibit the proliferation of medullary thyroid cancer cells in a time and dose dependent. When parathyroid hormone concentra tion reached a concentration of 2.0 IJmol/L, the parathyroid hormone receptor monoclonal antibody reached a concentration of 1.0 μmol/L, the cell growth was most significantly inhibited （P 〈 0.05）. Conclusion： Parathyroid hormone and parathyroid hormone receptor monoclonal antibody were able to inhibit the proliferation of medullary thyroid carcinoma cells and signifi cantly reduce the proliferation index.

Single nucleus/cell RNA-seq of the chicken hypothalamicpituitaryovarian axis offers new insights into the molecular regulatory mechanisms of ovarian development

The hypothalamic-pituitary-ovarian(HPO)axis represents a central neuroendocrine network essential for reproductive function.Despite its critical role,the intrinsic heterogeneity within the HPO axis across vertebrates and the complex intercellular interactions remain poorly defined.This study provides the first comprehensive,unbiased,cell type-specific molecular profiling of all three components of the HPO axis in adult Lohmann layers and Liangshan Yanying chickens.Within the hypothalamus,pituitary,and ovary,seven,12,and 13 distinct cell types were identified,respectively.Results indicated that the pituitary adenylate cyclase activating polypeptide(PACAP),follicle-stimulating hormone(FSH),and prolactin(PRL)signaling pathways may modulate the synthesis and secretion of gonadotropin-releasing hormone(GnRH),FSH,and luteinizing hormone(LH)within the hypothalamus and pituitary.In the ovary,interactions between granulosa cells and oocytes involved the KIT,CD99,LIFR,FN1,and ANGPTL signaling pathways,which collectively regulate follicular maturation.The SEMA4 signaling pathway emerged as a critical mediator across all three tissues of the HPO axis.Additionally,gene expression analysis revealed that relaxin 3(RLN3),gastrin-releasing peptide(GRP),and cocaine-and amphetamine regulated transcripts(CART,also known as CARTPT)may function as novel endocrine hormones,influencing the HPO axis through autocrine,paracrine,and endocrine pathways.Comparative analyses between Lohmann layers and Liangshan Yanying chickens demonstrated higher expression levels of GRP,RLN3,CARTPT,LHCGR,FSHR,and GRPR in the ovaries of Lohmann layers,potentially contributing to their superior reproductive performance.In conclusion,this study provides a detailed molecular characterization of the HPO axis,offering novel insights into the regulatory mechanisms underlying reproductive biology.

Evaluation of steroidogenic capacity after follicle stimulating hormone stimulation in bovine granulosa cells of Revalor 200~ implanted heifers

Background： Heifers not used as breeding stock are often implanted with steroids to increase growth efficiency thereby altering hormone profiles and potentially changing the environment in which ovarian follicles develop. Because bovine granulosa cell culture is a commonly used technique and often bovine ovaries are collected from abattoirs with no record of implant status, the objective of this study was to determine if the presence of an implant during bovine granulosa cell development impacts follicle stimulating hormone-regulated steroidogenic enzyme expression. Paired ovaries were collected from 16 feedlot heifers subjected to 1 of 3 treatments： non-implanted （n = 5）, Revalor 200 for 28 d （n = 5）, or Revalor 200 for 84 d （n = 6）. Small follicle （1 to 5 mm） granulosa cells were isolated from each pair and incubated with phosphate buffered saline （n = 16） or 100 ng/mL follicle stimulating hormone （n = 16） for 24 h. Results： Granulosa cells of implanted heifers treated with follicle stimulating hormone produced medium concentrations of progesterone similar （P = 0.22） to non-implanted heifers, while medium estradiol concentrations were increased （P 〈 0.10） at 28 and 84 d compared to non-implanted heifers indicating efficacy of treatment. Additionally, real-time PCR analysis in response to follicle stimulating hormone treatment demonstrated a decrease in steroidogenic acute regulatory protein （P = 0.05） mRNA expression in heifers implanted for 84 d and an increase in P450 side chain cleavage mRNA in granulosa cells of heifers implanted for 28 （P 〈 0.10） or 84 d （P 〈 0.05） compared to non-implanted females. However, no difference in expression of 3-beta-hydroxysteroid dehydrogenase （P= 0.57） and aromatase （P = 0.23） were demonstrated in implanted or non-implanted heifers. Conclusions： These results indicate follicles which develop in the presence of high concentrations of androgenic and estrogenic steroids via an implant tend to demonstrate an altered capacity to respond to follicle stimulating hormone stimulation. Thus, efforts should be made to avoid the use of implanted heifers to study steroidogenesis in small follicle granulosa cell culture systems.

Small cell lung cancer with panhypopituitarism due to ectopic adrenocorticotropic hormone syndrome: A case report

BACKGROUND Small cell lung cancer(SCLC)accounts for 15%of lung cancers,and it commonly expresses peptide and protein factors that are active as hormones.These secreting factors manifest as paraneoplastic disorders,such as ectopic adrenocorticotropic hormone(ACTH)syndrome(EAS).The clinical features are abnormalities in carbohydrate metabolism,hypokalemia,peripheral edema,proximal myopathy,hypertension,hyperpigmentation,and severe systemic infection.However,it is uncommon that EAS has an influence on hypothalamus-pituitary function.CASE SUMMARY A 62-year-old man presented with complaints of haemoptysis,polyuria,polydipsia,increased appetite,weight loss,and pigmentation.Following a series of laboratory and imaging examinations,he was diagnosed with SCLC,EAS,hypogonadism,hypothyroidism,and central diabetes insipidus.After three rounds of chemotherapy,levels of ACTH,cortisol,thyroid hormone,gonadal hormone,and urine volume had returned to normal levels.In addition,the pulmonary tumor was reduced in size.CONCLUSION We report a rare case of SCLC complicated with panhypopituitarism due to EAS.We hypothesize that EAS induced high levels of serum glucocorticoid and negative feedback for the synthesis and secretion of antidiuretic hormone from the paraventricular nucleus,and trophic hormones from the anterior pituitary.Therefore,patients who present with symptoms of hypopituitarism,or even panhypopituitarism,with SCLC should be evaluated for EAS.

Stability of Human Follicle-Stimulating Hormone Receptor mRNA in Stably Transfected Cells

In order to assess the impact of mRNA degradation on steady state levels of follicle stimulating hormone receptor (FSHR) mRNA and on regulation of FSHR gene expression, the stability and half life of FSHR mRNA were determined in transfected cells expressing recombinant FSHR. Time dependent changes in FSHR mRNA content were determined by nuclease protection solution hybridization assay (NPA) or by qualitative reverse transcription competitive polymerase chain reaction (RT PCR) in cultured hFSHR YI cells, cell lines stably transfected with a human FSHR cDNA. FSHR mRNA content remained constant during 8 h control incubations of hFSHR Y1 cells (NPA, 2.9±0.3 μg/mg RNA; RT PCR, 2.7±0.3 μg/mg RNA). Actinomycin D (ActD, 5 μg/ml) inhibited mRNA synthesis, as assessed by incorporation of uridine into total RNA, by 90 % within 1 h in hFSHR Y1 cells. No effect of ActD on cellular morphology or viability was observed. ActD caused a time dependent decrease in FSHR mRNA content in hFSHR Y1 cell lines with a lag time of 1 h. There were no significant differences in the rate of FSHR mRNA degradation between the two methods of mRNA quantification. The half life of hFSHR mRNA was 3.6±0.2 h by NPA and 3.1±0.1 h by RT PCR. The results indicated that degradation of mRNA was an important process in maintenance of steady state expression of the FSHR gene in cells stably expressing recombinant receptor.

Follicle-stimulating Hormone(FSH) Induced Internalization of Porcine FSH Receptor in Cultured Porcine Granulosa Cells and Chinese Hamster Ovary Cells Transfected with Recombinant Porcine FSH Receptor cDNA

In order to study the fate of human follicle-stimulating hormone (FSH) when hormone binds to its receptor, a quick biochemical method that can differentiate between the surface-bound and internalized hormone was used to determine the internalization induced by FSH in cultured both porcine granulosa cells and Chinese hamster ovary (CHO) cells expressing recombinant porcine FSH receptor. The results showed that FSH was slowly internalized, and the internalized radioactivity (acid resistant) reached a peak 10-12 h after addition of 125 I-hFSH. It was suggested that FSHR do not get internalized rapidly under physiological circumstances precisely because the appropriate sequences are absent.

Effects of Antisense Oligodeoxynucleotide to Follicle-stimulating Hormone Receptor on the Cell Proliferation and Apoptosis in Cells Derived from Human Ovarian Mucinous Cystadenocarcinoma in Vitro

The human ovarian mucinous cystadenocarcinoma （hOMC） cells were co-cultured with antisense oligodeoxynucleotide （antisense ODN）, nonsense ODN, and follicle-stimulating hormone （FSH） at different concentrations for the purpose of observing the effects of antisense ODN to FSH receptor （FSHR） on the proliferation and apoptosis of cultured hOMC cells in vitro. The inhibitory rates of growth were measured by using MTT method on the 2nd, 4th, 6th, 8th and 10th days after the interference of antisense ODN, nonsense ODN, and FSH, respectively. The apoptotic rates and the cell cycles were determined by means of flow cytometry, the apoptosis indexes were detected by using TUNEL, and the expression of caspase-3 was measured by using SP immunohistochemistry. Compared with that in the control group, the proliferative activity of hOMC cells was increased obviously in FSH groups （P〈0.05 or P〈0.01）, decreased distinctly in antisense ODN groups （P〈0.05 or P〈0.01）, and unchanged in nonsense ODN groups, respectively. Meanwhile, antisense ODN could significantly antagonize the FSH-promoted cell proliferative activity （P〈0.01）. Compared with those in the control group, the apoptotic rates and the expression of caspase-3 were dramatically increased in the mid- and high-dose antisense ODN groups （P〈0.05 or P〈0.01）, while the number of cells in G1/G0 phase was significantly decreased and that in S phase distinctly increased （P〈0.01）, There was no change in nonsense ODN groups （P〉0.05）, It was suggested that FSH may improve the development of hOMC cells, However, antisense ODN could inhibit proliferative activity and the FSH-promoted proliferative activity in hOMC cells, at the same time, antisense ODN could inhibit hOMC cell growth by inducing apoptosis.